Our recently published studies indicate that regulatory T cells (Tregs) expand in response to Mycobacterium tuberculosis (M. tb) in healthy tuberculin reactors and some CD4?Foxp3- cells are converted to CD4???? cells. Limited information is available about the cellular and molecular mechanism that expand Tregs during M. tb infection. Our preliminary data suggest that expression of PD-1, IL-27 and cytokine inducible SH2-containing protein (CISH) control expansion of M. tb-induced Tregs. We also made the surprising observation that that M. tb-expanded Tregs inhibit replication of M. tb and induce expression of the M. tb dosR gene, which regulates multiple genes that are expressed when M. tb enters a non-replicative persistent state. Furthermore, we made the unexpected observation that natural killer (NK) cells regulate M. tb-expanded Tregs by lysing them. We propose the following aims. 1) Determine the molecular mechanisms through which PD-1 and IL-27 induce Treg expansion in response to M. tb. We will first determine if PD-1 and IL-27 induce CISH expression. Next, we will delineate the signaling pathways and transcription factors downstream of CISH that enhance FoxP3 expression 2) Determine the mechanisms by which Tregs inhibit mycobacterial growth and induce expression of dosR by M. tb. We will determine if the upregulation of dosR and genes of the dosR regulon cause the reduced intracellular growth mediated by Tregs. Next, we will determine if Treg-mediated dosR expression and intracellular growth inhibition are mediated through heme oxygenase, and/or through nitric oxygenase production by macrophages. 3) Determine the mechanisms by which NK cells inhibit expansion of Tregs. We will identify the NK cell subpopulations that lyse M. tb-expanded Tregs and determine the cellular and molecular mechanisms that upregulate ULBP1 expression in activated Tregs. We will also determine if NK cells lyse expanded Tregs in mice in vivo. PUBLIC HEALTH RELEVANCE: Tuberculosis causes almost 2 million deaths per year. A subpopulation of blood cells called Tregulatory cells (Tregs) inhibit the immune response to tuberculosis . We will determine the mechanisms by which Tregs expand and are eliminated from the body. This work will lead to development of better methods to treat and prevent tuberculosis.